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Non-LTR Retrotransposons in Mosquitoes: Diversity, Evolution, and Analysis of Potentially Active ElementsBiedler, James K. 23 August 2005 (has links)
This research focuses on non-Long Terminal Repeat (non-LTR) retrotransposons in the African malaria mosquito, Anopheles gambiae and other mosquito species. An unprecedented diversity of non-LTRs was discovered by genome analysis of the An. gambiae genome assembly. One hundred and four families were found by a reiterative and comprehensive search using the conserved reverse transcriptase domains of known non-LTRs from a number of organisms as the starting queries. These families range in copy number from a few to approximately 2000 and occupy at least 3% of the genome. An. gambiae non-LTRs represent 8 of the 15 previously defined clades, plus two novel clades, Loner and Outcast, raising the total number of known clades to 17. The first invertebrate L1 clade representatives were also found. All clades except one have families with sequence characteristics suggesting recent activity.
Juan, a non-LTR of the Jockey clade originally discovered in the mosquito Culex pipiens quinquefasciatus (Mouches et al. 1991), has been implicated in horizontal transfer in three non-sibling species of the Aedes genus (Mouches, Bensaadi, and Salvado 1992). PCR was used to obtain sequences from 18 mosquito species of six genera. Phylogenetic analysis demonstrates predominant vertical inheritance of Juan elements among these species. There is strong evidence from sequence analysis supporting the recent activity of Juan in several divergent species. We hypothesize that the sustained activity (versus quick inactivation) of non-LTRs in mosquitoes may contribute to the diversity we observe in the An. gambiae genome today.
Promoter and transcriptional analyses were performed for several families previously identified as potentially active elements based on sequence analysis. RT-PCR results indicate that transcripts are present in An. gambiae cell lines that contain sequences corresponding to 13 of 15 tested non-LTR families. The 5' UTRs of An. gambiae non-LTRs from the I, Jockey, and L1 clades support basal transcription in divergent mosquito cell lines from 3 species. The Jen-1 5'UTR did not support transcription in Ae. aegypti and had low activity in Ae. albopictus. In summary, this research shows that Non-LTRs have been highly successful genomic elements that have flourished in many divergent mosquito species. / Ph. D.
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Dynamique et évolution de deux lignées remarquables de rétrotransposons à LTR dans le genre Coffea (famille des Rubiacées) / Dynamic dans evolution of two notable LTR retrotransposons lineages in Coffea genus (Rubiaceae family)Dupeyron, Mathilde 23 November 2017 (has links)
Les éléments transposables (ET) sont des portions d’ADN capables de se déplacer et d’augmenter le nombre de leurs copies dans les génomes. Deux grands types de transposition, correspondant à deux grandes classes d’ET, sont retrouvés chez la quasi-totalité des génomes étudiés à ce jour. Les rétrotransposons à LTR (Long Terminal Repeats, LTR-RT), appartenant à la Classe 1, sont les composants majoritaires des génomes des plantes. Leur prolifération peut avoir un impact important sur l’organisation, la variation de taille, l’évolution des génomes et l’activité des gènes.Le café, largement consommé dans le monde et produit uniquement par des pays du Sud, est issu de deux espèces cultivées d’origine africaine : Coffea arabica et C. canephora. Le genre Coffea est constitué de 139 espèces occupant des habitats très variés en Afrique, dans les îles de l’ouest de l’océan Indien, l’Inde, l’Asie tropicale et du sud-est et au nord de l’Australie. Toutes les espèces son diploïdes, à l’exception notable de C. arabica, allotétraploïde, issu d’une hybridation interspécifique récente entre les deux espèces diploïdes : C. canephora et C. eugenioides. Pour autant, la taille des génomes des espèces diploïdes varie du simple au double. Les nombreuses données génomiques aujourd’hui disponibles au sein du genre Coffea permettent d’étudier la dynamique des LTR-RT constituant au minimum 42% du génome de C. canephora, l’espèce séquencée et disponible dans les bases de données publiques.Dans ce travail, deux lignées remarquables de LTR-RT, Bianca et SIRE, ont été étudiées par des approches bio- informatiques. Bianca sensu stricto, présente uniquement chez les monocotylédones, est représentée chez les dicotylédones par la famille Divo, très peu étudiée à ce jour. L’activation récente de Divo sans induire sa propre structuration, est étroitement associée à la différenciation génétique de C. canephora. Par contre, tout en étant présente dans toutes les espèces de caféiers étudiées, l’activation semble sporadique. À l’opposé, les éléments SIRE, la seule lignée de LTR-RT de la superfamille des Copia contenant un domaine enveloppe comme les rétrovirus, montre des variations structurales importantes entre les accessions des espèces diploïdes à l’origine de C arabica et plus globalement, et en parallèle de l’évolution du genre.Nos travaux montrent que la compréhension de la dynamique des LTR-RT dans un genre peut permettre de mieux appréhender son histoire évolutive, chaque famille de LTR-RT pouvant apporter un éclairage différent. Nos résultats indiquent qu’à la fois les clades biogéographiques (phylogénie moléculaire des caféiers) mais aussi certaines accessions d’espèces diploïdes ont des histoires particulières. Celles-ci seraient vraisemblablement liées à la colonisation de nouvelles niches et à la dynamique des LTR-RT composant les génomes des Coffea. / Transposable elements (TEs) are DNA fragments that are able to move and to increase their copy numbers. Two transposition mechanisms corresponding to the two main TE classes are found in almost all organisms. LTR retrotransposons (Long Terminal Repeats, LTR-RTs), belonging to Class 1, are the main components of plant genomes. Genome organisation, size variation, evolution and gene activity can be strongly impacted by their proliferation.Worldwide consumed and produced by South countries, coffee is obtained from two African cultivated species: Coffea arabica and C. canephora. The Coffea genus includes 139 species occurring in diverse habitats in Africa, Madagascar, Mascarene Islands, Comoros, India, Southeast and Tropical Asia and North Australia. All the species are diploids, except the noteworthy allotetraploid C. arabica, originated from a recent inter-specific hybridisation between two diploids: C. canephora and C. eugenioides. However, genome size of diploid species can vary for up to two folds. Today, the numerous genomic data available for Coffea allows the study of LTR- RTs, constituting at least 42% of C. canephora genome, the sequenced species available in public databases.In this work, two notable LTR-RT lineages, Bianca and SIRE, have been studied by bioinformatics approaches. Bianca s.s., is present only in Monocots and it is represented in Dicots by the Divo family, poorly studied nowadays. The recent activation of Divo, without leading to its own structuring, is closely associated to the genetic differentiation of C. canephora. However, this activation seems sporadic as being present in all the coffee-trees species studied here. On the opposite, SIRE elements, which are the only Copia LTR-RTs carrying an envelope-like gene as retroviruses, show an important structuring variation between accessions among C. arabica progenitors, and in parallel to the genus evolution.Our work shows that understanding the LTR-RTs dynamics in a genus allows a better perception of its evolutionary history, with the possibility of different evolutionary timing given by different LTR-RTs families. Our results also indicate that both the biogeographic clades (coffee molecular phylogeny) and also some diploid accessions have peculiar histories, probably related to the colonisation of new ecological niches and to the LTR- RTs dynamics.
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Structural and functional characterization of giant plant Ogre-like retrotransposons / Structural and functional characterization of giant plant Ogre-like retrotransposonsSTEINBAUEROVÁ, Veronika January 2012 (has links)
Ogre elements represent a distinct group of Ty3/gypsy LTR retrotransposons occurring in a range of dicot plants. They are characterized by two specific features ? presence of long extra open reading frame in 5´ untranslated region with unknown function and a non-coding sequence containing several stop codons separating protease and reverse transcriptase domains which was proposed to be removed by splicing. This thesis describes the functional analysis of intron splicing in Ogre retrotransposons. Further, it investigates additional coding information not only in Ogre retrotransposons but in the whole group of Ty3/gypsy retroelements.
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Bioinformatický nástroj pro anotaci transposonů / Bioinformatics Tool for Transposons AnnotationJenčo, Michal January 2017 (has links)
This thesis provides theoretical resources for the design of a new bioinformatics tool for transposon annotation with focus on their additional structural elements. There is a biological description of transposons, the mobile elements in DNA, their classification and structure. It further deals with the overview and classification of available transposon identification and annotation bioinformatics tools, description of function and implementation of a select few. Next we state the scheme of a new bioinformatics tool for LTR retrotransposon identification and annotation with a focus on extra ORFs and tandem repeats. The functionality of this new tool was tested on the A. thaliana genome. We identified 95 groups of conserved extra ORFs and 10 groups of conserved tandem repeats.
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Análise transcriptômica de genes e LTR retrotransposons em arroz (Oryza sativa ssp. japonica) em resposta à toxidez por ferro / Transcriptomic analysis of genes and LTR retrotransposons in rice (Oryza sativa ssp. japonica) in response to iron toxicityFinatto, Taciane 27 February 2012 (has links)
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Previous issue date: 2012-02-27 / Iron toxicity in plants is associated with the presence of large concentrations of reduced iron (Fe2+) in the soil solution, which occurs in flooded soils and affects rice plants grown under this condition. Symptoms of iron toxicity involve oxidative stress in leaves, as a response to excessive Fe2+ absorption by the roots. The responses of plants to stress conditions include stimulus perception, signal transduction and gene transcription activation. Besides gene expression, LTR (Long Terminal Repeat) retrotransposons represent ca. 22% of the rice genome, they can be transcriptionally activated under stress, and they can alter the expression of adjacent genes (e.g. due to alterations in chromatin structure). This study aimed to identify differentially expressed genes and LTR retrotransposons in leaves of 18-day-old rice seedlings (Oryza sativa ssp. japonica cv. Nipponbare) after four days of iron excess exposure. They were identified a differential expression of genes and LTR retrotransposons in rice exposed to iron excess using a microarray approach. Total RNA was extracted from leaves of 18-day-old rice seedlings (Oryza sativa L. ssp japonica cv. Nipponbare) after four days of cultivation in nutrient solution with iron excess (7 mM of FeSO47H2O) and in a control solution. The hybridization was performed with cDNA and rice transposome array v. 2.0 microarray (Roche/NimbleGen technology, an improvement of v.1.0, Picault et al., 2009). Data from gene expression was analyzed by the Bayesian t-test with BH adjustment method. Gene annotation, gene ontology, and LTR retrotransposon identification were performed at RAP-DB (Rice Annotation Project Database, build 5), and microarray results were validated by RT-qPCR. Considering log2 FC (log2-fold-change) ≤ -1 as underexpression and ≥ 1 as overexpression (p-values ≤ 0.05), 44 down-regulated and 1,572 up-regulated genes with described function were identified. Down-regulated genes were related to a wide range of functions and no gene family could be highlighted. Among the up-regulated genes, 166 were transcription factors, the most representative belonging to the Zinc finger RING/FYVE/PHD-type family (22) and WRKY family (19); other genes were from the kinase family, participating in biological processes of protein amino acid phosphorylation (86); had molecular function of iron ion binding (56); were involved in response to oxidative stress (scavenging of reactive oxygen species) (26); had molecular function of transport activity (84), including four genes related to heavy metal transport/detoxification and four genes of the multi antimicrobial extrusion protein MATE family; and were involved in the biological process of apoptosis (14), including 10 genes of NB-ARC. Among the up-regulated genes, 435 present at least one cis-regulatory element responsive to abscisic acid (ABA) with significant occurrence (P≤0.05) in its promoter region (1 kbp upstream of the transcription start site). These data indicate that about 28% of the up-regulated genes can be regulated by changing in the ABA content in leaves in response to iron excess. Regarding expression of LTR retrotransposons, 302 were down-regulated (53 Ty1/Copia, 172 Ty3/Gypsy and 77 unclassified), and 4342 up-regulated (466 Ty1/Copia, 2276 Ty3/Gypsy and 1600 unclassified). They were observed a large activity of LTR retrotransposons in response to iron toxicity, and furthermore, they were verified that LTR retrotransposons transcription can extend to 5' and 3' flanking regions. In addition, 16 situations that should up-regulated LTR retrotransposons are located at a very short distance (smaller than 1000 base pairs) in the same chromosome of up-regulated genes suggesting co-transcription, these occurrences are represented by eight where the LTR retrotransposon and the gene have the same sense of transcription (plus); five occurrences with the both with the same sense of transcription (minus) and one occurrence where they have opposite senses. Additionally, two occurrences that in which both, DNA sequences of up-regulated retrotransposon and gene, are overlapped and have the same sense of transcription. / A toxidez por ferro em plantas está associada com a presença de grandes concentrações de ferro (Fe) reduzido (Fe2+) na solução do solo, esta condição pode ocorrer em solos irrigados por inundação. Os sintomas de toxidez por ferro incluem estresse oxidativo nas folhas como resultado do excesso de Fe2+ absorvido pelas raízes, resultando em perdas na produtividade. As respostas das plantas às condições de estresse envolvem a percepção dos estímulos, transdução de sinais e ativação da transcrição gênica. Além da expressão gênica, os LTR retrotransposons (Long Terminal Repeat Retrotransposons) que respresentam cerca de 20% do genoma do arroz, podem ser transcricionalmente ativados em condições de estresse e desta forma, influenciar a expressão de genes adjacentes (por exemplo devido a alterações na estrutura da cromatina). Este estudo teve por objetivo identificar genes e LTR retrotransposons diferencialmente expressos em plântulas de arroz (Oryza sativa ssp. japonica cv. Nipponbare), após quatro dias de exposição ao excesso de ferro em solução nutritiva. A expressão diferencial de genes e LTR retrotransposons foi analisada utilizando a técnica de microarranjo e sua validação foi realizada por meio de RT-qPCR. O RNA total foi extraído de folhas de plântulas de arroz cv. Nipponbare, após quatro dias de cultivo em solução nutritiva adicionada de ferro na concentração de 7 mM (FeSO47H2O) (presença de toxidez) e a condição controle com presença de ferro na concentração de 10 μM. O cDNA fita dupla foi sintetitizado a partir do RNA mensageiro. A hibridização foi realizada entre o cDNA das duas condições em triplicatas biológicas e o microarranjo Rice Transposome Array v. 2.0 (Roche/NimbleGen technology, an improvement of v.1.0, Picault et al., 2009). Os valores de intensidade de cada spot foram normalizados, transformados e comparados pelo teste T Bayesiano. A identificação dos genes e LTR retrotransposons foi realizada de acordo com o banco de dados RAP-DB (Rice Annotation Project Database, build 5). Considerando log2 FC (log2-fold-change) ≤ -1 como subexpressão e ≥ 1 como superexpressão e P≤ 0.05 para ambas condições. Foram identificados 44 genes subexpressos e 1.572 superexpressos com funções descritas. Os genes subexpressos desempenham a uma vasta gama de funções. Entre elas destacam-se: 166 genes que são fatores de transcrição, sendo que os mais representativos pertencem à família Zinc finger RING/FYVE/PHD-type family (22 genes) e WRKY (19 genes); outros genes da família das cinases que participam também da sinalização celular em processos biológicos de fosforilação de aminoácidos nas proteínas (86 genes); outros genes com função molecular de ligação ao íon ferro (56 genes); 26 genes envolvidos na resposta ao estresse oxidativo (scavengers de espécies reativas de oxigênio); 84 genes com função molecular de transporte, incluindo quatro genes relacionados ao transporte e detoxificação de metais pesados e quatro genes da família MATE; 14 genes envolvidos em apoptose, incluindo 10 genes NB-ARC. Entre os genes superexpressos, 435 apresentam pelo menos um elemento regulatório de ação cis responsivo ao ácido abscisico (ABA) com ocorrência significativa (P≤0,05) em sua região promotora (1 kbp a montante do sítio de início da transcrição). Estes dados indicam que cerca de 28% dos genes superexpressos podem ser regulados pelas alterações no conteúdo de ABA nas folhas, em resposta ao estresse por excesso de ferro. Considerando a expressão do LTR retrotransposons, 302 apresentaram subexpressão (53 Ty1/Copia, 172 Ty3/Gypsy e 77 não classificados), e 4.342 apresentaram superexpressão (466 Ty1/Copia, 2276 Ty3/Gypsy e 1600 não classificados). Foi constatada grande atividade transcricional dos LTR retrotransposons em resposta à toxidez por ferro, sendo que a transcrição dos LTR retrotransposons pode se estender às suas regiões flanqueadoras 5 e 3 , além disso foram encontradas 16 ocorrencias em que o LTR retrotransposon e o gene superexpresso estão localizados a uma distância menor do que 1000 pares de bases no mesmo cromossomo, sugerindo co-transcrição entre ambos. Entre as 16 ocorrências, oito em que o LTR retrotransposon e o gene apresentam o mesmo sentido de transcrição (plus); cinco ocorrências com mesmo sentido de transcrição (minus) e uma ocorrência onde LTR retrotrotransposon e gene apresentam sentidos de transcrição opostos. Foram observadas ainda, duas ocorrências em que as sequencias de DNA do LTR retrotransposon e do gene superexpressos estão sobrepostas, e apresentam o mesmo sentido de transcrição.
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Análise transcriptômica de genes e LTR retrotransposons em arroz (Oryza sativa ssp. japonica) em resposta à toxidez por ferro / Transcriptomic analysis of genes and LTR retrotransposons in rice (Oryza sativa ssp. japonica) in response to iron toxicityFinatto, Taciane 27 February 2012 (has links)
Made available in DSpace on 2014-08-20T13:25:37Z (GMT). No. of bitstreams: 1
tese_taciane_finatto.pdf: 5834731 bytes, checksum: e10f781234d54582cc17a9b8dff16c53 (MD5)
Previous issue date: 2012-02-27 / Iron toxicity in plants is associated with the presence of large concentrations of reduced iron (Fe2+) in the soil solution, which occurs in flooded soils and affects rice plants grown under this condition. Symptoms of iron toxicity involve oxidative stress in leaves, as a response to excessive Fe2+ absorption by the roots. The responses of plants to stress conditions include stimulus perception, signal transduction and gene transcription activation. Besides gene expression, LTR (Long Terminal Repeat) retrotransposons represent ca. 22% of the rice genome, they can be transcriptionally activated under stress, and they can alter the expression of adjacent genes (e.g. due to alterations in chromatin structure). This study aimed to identify differentially expressed genes and LTR retrotransposons in leaves of 18-day-old rice seedlings (Oryza sativa ssp. japonica cv. Nipponbare) after four days of iron excess exposure. They were identified a differential expression of genes and LTR retrotransposons in rice exposed to iron excess using a microarray approach. Total RNA was extracted from leaves of 18-day-old rice seedlings (Oryza sativa L. ssp japonica cv. Nipponbare) after four days of cultivation in nutrient solution with iron excess (7 mM of FeSO47H2O) and in a control solution. The hybridization was performed with cDNA and rice transposome array v. 2.0 microarray (Roche/NimbleGen technology, an improvement of v.1.0, Picault et al., 2009). Data from gene expression was analyzed by the Bayesian t-test with BH adjustment method. Gene annotation, gene ontology, and LTR retrotransposon identification were performed at RAP-DB (Rice Annotation Project Database, build 5), and microarray results were validated by RT-qPCR. Considering log2 FC (log2-fold-change) ≤ -1 as underexpression and ≥ 1 as overexpression (p-values ≤ 0.05), 44 down-regulated and 1,572 up-regulated genes with described function were identified. Down-regulated genes were related to a wide range of functions and no gene family could be highlighted. Among the up-regulated genes, 166 were transcription factors, the most representative belonging to the Zinc finger RING/FYVE/PHD-type family (22) and WRKY family (19); other genes were from the kinase family, participating in biological processes of protein amino acid phosphorylation (86); had molecular function of iron ion binding (56); were involved in response to oxidative stress (scavenging of reactive oxygen species) (26); had molecular function of transport activity (84), including four genes related to heavy metal transport/detoxification and four genes of the multi antimicrobial extrusion protein MATE family; and were involved in the biological process of apoptosis (14), including 10 genes of NB-ARC. Among the up-regulated genes, 435 present at least one cis-regulatory element responsive to abscisic acid (ABA) with significant occurrence (P≤0.05) in its promoter region (1 kbp upstream of the transcription start site). These data indicate that about 28% of the up-regulated genes can be regulated by changing in the ABA content in leaves in response to iron excess. Regarding expression of LTR retrotransposons, 302 were down-regulated (53 Ty1/Copia, 172 Ty3/Gypsy and 77 unclassified), and 4342 up-regulated (466 Ty1/Copia, 2276 Ty3/Gypsy and 1600 unclassified). They were observed a large activity of LTR retrotransposons in response to iron toxicity, and furthermore, they were verified that LTR retrotransposons transcription can extend to 5' and 3' flanking regions. In addition, 16 situations that should up-regulated LTR retrotransposons are located at a very short distance (smaller than 1000 base pairs) in the same chromosome of up-regulated genes suggesting co-transcription, these occurrences are represented by eight where the LTR retrotransposon and the gene have the same sense of transcription (plus); five occurrences with the both with the same sense of transcription (minus) and one occurrence where they have opposite senses. Additionally, two occurrences that in which both, DNA sequences of up-regulated retrotransposon and gene, are overlapped and have the same sense of transcription. / A toxidez por ferro em plantas está associada com a presença de grandes concentrações de ferro (Fe) reduzido (Fe2+) na solução do solo, esta condição pode ocorrer em solos irrigados por inundação. Os sintomas de toxidez por ferro incluem estresse oxidativo nas folhas como resultado do excesso de Fe2+ absorvido pelas raízes, resultando em perdas na produtividade. As respostas das plantas às condições de estresse envolvem a percepção dos estímulos, transdução de sinais e ativação da transcrição gênica. Além da expressão gênica, os LTR retrotransposons (Long Terminal Repeat Retrotransposons) que respresentam cerca de 20% do genoma do arroz, podem ser transcricionalmente ativados em condições de estresse e desta forma, influenciar a expressão de genes adjacentes (por exemplo devido a alterações na estrutura da cromatina). Este estudo teve por objetivo identificar genes e LTR retrotransposons diferencialmente expressos em plântulas de arroz (Oryza sativa ssp. japonica cv. Nipponbare), após quatro dias de exposição ao excesso de ferro em solução nutritiva. A expressão diferencial de genes e LTR retrotransposons foi analisada utilizando a técnica de microarranjo e sua validação foi realizada por meio de RT-qPCR. O RNA total foi extraído de folhas de plântulas de arroz cv. Nipponbare, após quatro dias de cultivo em solução nutritiva adicionada de ferro na concentração de 7 mM (FeSO47H2O) (presença de toxidez) e a condição controle com presença de ferro na concentração de 10 μM. O cDNA fita dupla foi sintetitizado a partir do RNA mensageiro. A hibridização foi realizada entre o cDNA das duas condições em triplicatas biológicas e o microarranjo Rice Transposome Array v. 2.0 (Roche/NimbleGen technology, an improvement of v.1.0, Picault et al., 2009). Os valores de intensidade de cada spot foram normalizados, transformados e comparados pelo teste T Bayesiano. A identificação dos genes e LTR retrotransposons foi realizada de acordo com o banco de dados RAP-DB (Rice Annotation Project Database, build 5). Considerando log2 FC (log2-fold-change) ≤ -1 como subexpressão e ≥ 1 como superexpressão e P≤ 0.05 para ambas condições. Foram identificados 44 genes subexpressos e 1.572 superexpressos com funções descritas. Os genes subexpressos desempenham a uma vasta gama de funções. Entre elas destacam-se: 166 genes que são fatores de transcrição, sendo que os mais representativos pertencem à família Zinc finger RING/FYVE/PHD-type family (22 genes) e WRKY (19 genes); outros genes da família das cinases que participam também da sinalização celular em processos biológicos de fosforilação de aminoácidos nas proteínas (86 genes); outros genes com função molecular de ligação ao íon ferro (56 genes); 26 genes envolvidos na resposta ao estresse oxidativo (scavengers de espécies reativas de oxigênio); 84 genes com função molecular de transporte, incluindo quatro genes relacionados ao transporte e detoxificação de metais pesados e quatro genes da família MATE; 14 genes envolvidos em apoptose, incluindo 10 genes NB-ARC. Entre os genes superexpressos, 435 apresentam pelo menos um elemento regulatório de ação cis responsivo ao ácido abscisico (ABA) com ocorrência significativa (P≤0,05) em sua região promotora (1 kbp a montante do sítio de início da transcrição). Estes dados indicam que cerca de 28% dos genes superexpressos podem ser regulados pelas alterações no conteúdo de ABA nas folhas, em resposta ao estresse por excesso de ferro. Considerando a expressão do LTR retrotransposons, 302 apresentaram subexpressão (53 Ty1/Copia, 172 Ty3/Gypsy e 77 não classificados), e 4.342 apresentaram superexpressão (466 Ty1/Copia, 2276 Ty3/Gypsy e 1600 não classificados). Foi constatada grande atividade transcricional dos LTR retrotransposons em resposta à toxidez por ferro, sendo que a transcrição dos LTR retrotransposons pode se estender às suas regiões flanqueadoras 5 e 3 , além disso foram encontradas 16 ocorrencias em que o LTR retrotransposon e o gene superexpresso estão localizados a uma distância menor do que 1000 pares de bases no mesmo cromossomo, sugerindo co-transcrição entre ambos. Entre as 16 ocorrências, oito em que o LTR retrotransposon e o gene apresentam o mesmo sentido de transcrição (plus); cinco ocorrências com mesmo sentido de transcrição (minus) e uma ocorrência onde LTR retrotrotransposon e gene apresentam sentidos de transcrição opostos. Foram observadas ainda, duas ocorrências em que as sequencias de DNA do LTR retrotransposon e do gene superexpressos estão sobrepostas, e apresentam o mesmo sentido de transcrição.estresse oxidativo (scavengers de espécies reativas de oxigênio); 84 genes com função molecular de transporte, incluindo quatro genes relacionados ao transporte e detoxificação de metais pesados e quatro genes da família MATE; 14 genes envolvidos em apoptose, incluindo 10 genes NB-ARC. Entre os genes superexpressos, 435 apresentam pelo menos um elemento regulatório de ação cis responsivo ao ácido abscisico (ABA) com ocorrência significativa (P≤0,05) em sua região promotora (1 kbp a montante do sítio de início da transcrição). Estes dados indicam que cerca de 28% dos genes superexpressos podem ser regulados pelas alterações no conteúdo de ABA nas folhas, em resposta ao estresse por excesso de ferro. Considerando a expressão do LTR retrotransposons, 302 apresentaram subexpressão (53 Ty1/Copia, 172 Ty3/Gypsy e 77 não classificados), e 4.342 apresentaram superexpressão (466 Ty1/Copia, 2276 Ty3/Gypsy e 1600 não classificados). Foi constatada grande atividade transcricional dos LTR retrotransposons em resposta à toxidez por ferro, sendo que a transcrição dos LTR retrotransposons pode se estender às suas regiões flanqueadoras 5 e 3 , além disso foram encontradas 16 ocorrencias em que o LTR retrotransposon e o gene superexpresso estão localizados a uma distância menor do que 1000 pares de bases no mesmo cromossomo, sugerindo co-transcrição entre ambos. Entre as 16 ocorrências, oito em que o LTR retrotransposon e o gene apresentam o mesmo sentido de transcrição (plus); cinco ocorrências com mesmo sentido de transcrição (minus) e uma ocorrência onde LTR retrotrotransposon e gene apresentam sentidos de transcrição opostos. Foram observadas ainda, duas ocorrências em que as sequencias de DNA do LTR retrotransposon e do gene superexpressos estão sobrepostas, e apresentam o mesmo sentido de transcrição.
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2A-induced ribosome stallingOdon, Valèrie M. N. January 2014 (has links)
Originally 2A was characterised in foot-and-mouth disease virus. Site directed mutagenesis identified a C-terminus consensus motif [D(V/I)ExNPGP] and it is proposed that 2A interacts with the exit tunnel of the ribosome in a way that a specific peptide bond is skipped between the last glycine of 2A and the proline of 2B, thus providing a discontinuity in translation, resulting in release of discrete proteins from one single ORF. 2A was also identified in other picornaviruses, positive, single and double-stranded RNA insect viruses and mammalian rotaviruses. A motif present at the C-terminus of the 2A oligopeptide [D(V/I)ExNPGP] is very highly, though not completely conserved . The sequence upstream of this motif shows, however, no apparent conservation between 2As of different viruses. In this study, extensive site-directed mutagenesis were performed on several 2A sequences and a series of ‘hybrid' 2As comprising different consensus motifs juxtaposed with different upstream contexts were created as part of a detailed analysis of the mechanism of 2A-mediated ribosome stalling. The results demonstrated that a minimal region of twenty to twenty-three amino acids interacts with the exit tunnel of the ribosome to bring about a pause in processivity, alter the peptidyl transferase centre geometry and restrict the ribosome A site via two distinctive stalling mechanisms. Other molecular analyses tested here will require further optimisations or alternative methods: a visual method to explore the dynamics of re-initiation of translation from proline codon, purification of the translation-regulating factors and structural resolution of 2A sequences. Previously, cellular 2As were identified in non-LTR retrotransposons of trypanosomes. It is reported here as part of two other cellular organisms Saccoglossus kowalevskii (acorn worm) and Branchiostoma floridae (amphioxus). In the acorn worm, the nucleotides sequences corresponding to 2A motifs were part of the untranslated genome. In amphioxus, three 2A elements were identified in hypothetical proteins, and at the N-terminus of twenty non-LTR retrotransposons.
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L'influence du contexte génomique sur la sélection du site d'intégration par les rétrotransposons humains L1 / Influence of the genomic context on integration site selection by human L1 retrotransposonsSultana, Tania 12 December 2016 (has links)
Les rétrotransposons L1 (Long INterspersed Element-1) sont des éléments génétiques mobiles dont l'activité contribue à la dynamique du génome humain par mutagenèse insertionnelle. Les conséquences génétiques et épigénétiques d'une nouvelle insertion, et la capacité d'un L1 à être remobilisé, sont directement liées au site d’intégration dans le génome. Aussi, l’analyse des sites d’intégration des L1s est capitale pour comprendre leur impact fonctionnel - voire pathogène -, en particulier lors de la tumorigenèse ou au cours du vieillissement, et l’évolution de notre génome. Dans ce but, nous avons induit de façon expérimentale la rétrotransposition d'un élément L1 actif plasmidique dans des cellules en culture. Puis, nous avons cartographié les insertions obtenues de novo dans le génome humain grâce à une méthode de séquençage à haut-débit, appelée ATLAS-seq. Finalement, les sites pré-intégratifs identifiés par cette approche ont été analysés en relation avec un grand jeu de données publiques regroupant les caractéristiques structurales, génétiques ou épigénétiques de ces loci. Ces expériences ont révélé que les éléments L1 s’intègrent préférentiellement dans des régions de la chromatine faiblement exprimées et renfermant des activateurs faibles. Nous avons aussi trouvé plusieurs positions chromosomiques qui constituent des points chauds d'intégrations récurrentes. Nos résultats indiquent que la distribution des insertions de L1 de novo n’est pas aléatoire, que ce soit à l’échelle chromosomique ou à plus petite échelle, et ouvrent la porte à l'identification des déterminants moléculaires qui contrôlent la distribution chromosomique des L1s dans notre génome / Retrotransposons are mobile genetic elements that employ an RNA intermediate and a reverse transcription step for their replication. Long INterspersed Elements-1 (LINE-1 or L1) form the only autonomously active retrotransposon family in humans. Although most copies are defective due to the accumulation of mutations, each individual genome contains an average of 100 retrotransposition-competent L1 copies, which contribute to the dynamics of contemporary human genomes. L1 integration sites in the host genome directly determine the genetic consequences of the integration and the fate of the integrated copy. Thus, where L1 integrates in the genome, and whether this process is random, is critical to our understanding of human genome evolution, somatic genome plasticity in cancer and aging, and host-parasite interactions. To characterize L1 insertion sites, rather than studying endogenous L1 which have been subjected to evolutionary selective pressure, we induced de novo L1 retrotransposition by transfecting a plasmid-borne active L1 element into HeLa S3 cells. Then, we mapped de novo insertions in the human genome at nucleotide resolution by a dedicated deep-sequencing approach, named ATLAS-seq. Finally, de novo insertions were examined for their proximity towards a large number of genomic features. We found that L1 preferentially integrates in the lowly-expressed and weak enhancer chromatin segments. We also detected several hotspots of recurrent L1 integration. Our results indicate that the distribution of de novo L1 insertions is non-random both at local and regional scales, and pave the way to identify potential cellular factors involved in the targeting of L1 insertions
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Diversity and Evolution of Short Interspersed Nuclear Elements (SINEs) in Angiosperm and Gymnosperm Species and their Application as molecular Markers for GenotypingKögler, Anja 08 September 2020 (has links)
Short interspersed nuclear elements (SINEs) are small non-autonomous and heterogeneous retrotransposons, widespread in animals and plants and usually differentially propagated in related species resulting in genome-specific copy numbers.
Within the monocots, the Poaceae (sweet grasses) is the largest and economically most important plant family. The distribution of 24 Poaceae SINE (PoaS) families, five of which showing a subfamily structure, was analyzed in five important cereals (Oryza sativa, Triticum aestivum, Hordeum vulgare, Sorghum bicolor, Zea mays), the energy crop Panicum virgatum and the model grass Brachypodium distachyon. The comparative investigation of SINE abundance and sequence diversity within Poaceae species provides insights into their species‐specific diversification and amplification. The PoaS families and subfamilies fall into two length and structural categories: simple SINEs of up to 180 bp and dimeric SINEs larger than 240 bp. Of 24 PoaS families, 20 are structurally related across species, in particular either in their 5′ or 3′ regions. Hence, reshuffling between SINEs, likely caused by nested insertions of full-lengh and truncated copies, is an important evolutionary mechanism of SINE formation. Most striking, the recently evolved homodimeric SINE family PoaS‐XIV occurs exclusively in wheat (T. aestivum) and consists of two tandemly arranged PoaS‐X.1 copies.
Exemplary for deciduous tree species, the evolutionary history of SINE populations was examined in six Salicaceae genomes (Populus deltoides, Populus euphratica, Populus tremula, Populus tremuloides, Populus trichocarpa, Salix purpurea). Four of eleven Salicaceae SINE (SaliS) families exhibit a subfamily organization. The SaliS families consist of two groups, differing in their phylogenetic distribution pattern, sequence similarity and 3’ end structure. These groups probably emerged at different evolutionary periods of time: during the ‘salicoid duplication’ (~ 65 million years ago) in the Salix-Populus progenitor, and during the separation of the genus Salix (~ 45 - 65 million years ago), respectively. Similar to the PoaS families, the majority of the 20 SaliS families and subfamilies share regions of sequence similarity, providing evidence for SINE emergence by reshuffling. Furthermore, they also contain an evolutionarily young dimeric SINE family (SaliS-V), amplified only in two poplar genomes. The special feature of the Salicaceae SINEs is the contrast of the conservation of 5’ start motifs across species and SINE families compared to the high variability of
3’ ends within the SINE families, differing in sequence and length, presumably resulting from mutations in the poly(A) tail as a possible route for SINE elongation. Periods of increased transpositional activity promote the dissemination of novel 3’ ends. Thereby, evolutionarily older motifs are displaced leading to various 3’ end subpopulations within the SaliS families. Opposed to the PoaS families with a largely equal ratio of poly(A) to poly(T) tail SINEs, the SaliS families are exclusively terminated by adenine stretches.
Among retrotransposon-based markers, SINEs are highly suitable for the development of molecular markers due to their unidirectional insertion and random distribution mainly in euchromatic genome regions, together with an easy and fast detection of the heterogeneous SINE families. As a prerequisite for the development of SINE-derived inter-SINE amplified polymorphism (ISAP) markers, 13 novel Theaceae SINE families (TheaS-I - TheaS-VII, TheaS-VIII.1 and TheaS-VIII.2, TheaS-IX - TheaS-XIII) were identified in the angiosperm tree species Camellia japonica. Moreover, six Pinaceae SINE families (PinS-I.1 and PinS-I.2, PinS-II – PinS-VI) were detected in the gymnosperm species Larix decidua. Compared to the SaliS and PoaS families, structural relationships are less frequent within the TheaS families and absent in the PinS families.
The ISAP analysis revealed the genetic identity of Europe’s oldest historical camellia (C. japonica) trees indicating their vegetative propagation from the same ancestor specimen, which was probably the first living camellia on European ground introduced to England within the 18th century. Historical sources locate the native origin of this ancestral camellia specimen either in the Chinese province Yunnan or at the Japanese Gotō Islands. Comparative ISAPs showed no accordance to the Gotō camellia sample pool and appropriate Chinese reference samples were not available. However, the initial experiments demonstrated the potential of ISAP to resolve variations among natural populations.
The ISAP application on angiosperm trees also concerned fast growing Populus clones grown in short rotation coppice plantations for energy production. The species-specific P. tremula ISAP primers might also be applied for the discrimination of hybrid poplar clones involving P. tremuloides genome
portions, since SINEs of these two species are highly related. However, due to lineage-specific SINE evolution during speciation, cross-species applications are generally only successful to limited extent. The analysis of poplar hybrids composed of P. maximowiczii with either P. trichocarpa or P. nigra based on P. tremula ISAP primers showed a strongly reduced resolution.
In forestry, hybrid larch (e.g. Larix × eurolepis) genotypes have to be selected from the offspring of Japanese (Larix kaempferi) and European larch (Larix decidua) crosses, as they exhibit superior growth rates compared to the parental species. Initial ISAP-based examinations of European larch genotypes provided less polymorphic banding patterns, probably resulting from general high levels of synteny and collinearities reported for gymnosperm species. Hence, the ISAP was combined with the AFLP technique to the novel marker system inter-SINE-restriction site amplified polymorphism (ISRAP). The amplicons originating from genomic regions between SINEs and EcoRI cleavage sites were visualized with the sensitive capillary gel electrophoresis. The ISRAP assays, based on EcoRI adapter primers combined with two different SINE-derived primers, resulted in a sufficient number of polymorphic peaks to distinguish the L. decidua genotypes investigated. Compared to ISAPs, the ISRAP approach provides the required resolution to differentiate highly similar larch genotypes.
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